A Quick Guide to the

Mutation1717-1G–›A

C F T R S C I E N C E

© 2016 Vertex Pharmaceuticals Incorporated | VXR-HQ-02-00045a(1) | 03/2016

Loss of CFTR activity is the underlying cause of cystic fibrosis (CF)1

• People with 2 CFTR mutations resulting in loss of CFTR activity generally have a CF phenotype, which may include1-3,6

– Elevated sweat chloride (>60 mmol/L)

– Pancreatic insufficiency

– CBAVD a

– Lung function decline over time

– Pseudomonas aeruginosa colonization

aCBAVD, congenital bilateral absence of the vas deferens.

References: 1. Davis PB et al. Am J Respir Crit Care Med. 1996;154(5):1229-1256. 2. Rowe SM et al. Proc Am Thorac Soc. 2007;4(4):387-398. 3. Zielenski J. Respiration. 2000;67(2):117-133. 4. Sheppard DN et al. Nature. 1993;362(6416):160-164. 5. Welsh MJ, Smith AE. Cell. 1993;73(7): 1251-1254. 6. Castellani C et al. J Cyst Fibros. 2008;7(3):179-196.

Some CFTR mutations result in little to no CFTR activity3-5

0% 100%

Total CFTR Activity

0% 100%

Total CFTR Activity

Some CFTR mutations result in residual or partial CFTR activity3-5

Spectrum of Phenotypes Associated With Total CFTR Activity1,2

Total CFTR Activity

% of Normal

No CF Disease

CFTR-relatedDisorders

Cystic Fibrosis

Individuals with 2 normal alleles and CFTR mutation carriers

Clinical entities associated with CFTR dysfunctionthat do not fulfill diagnostic criteria for CF, e.g., CBAVD, acute recurrent or chronic pancreatitis and bronchiectasis

Typical phenotype: sinopulmonary disease, sweatchloride >60 mmol/L, pancreatic insufficiency, appearing early in life. In some patients, progressionof disease is delayed or not all features are present

100%

0%

Clinical Phenotype

2

Levels of CFTR activity affect survival in CF1

References: 1. McKone EF et al. Chest. 2006;130(5):1441-1447. 2. The World Bank. http://data.worldbank.org/indicator/SP.DYN.LE00.IN?page=2. Accessed November 12, 2015. 3. MacKenzie T et al. Ann Int Med. 2014;161:233-241.

1.00

0.75

0.50

0.25

0.00

0 10 20

Risk Time (Age)

Surv

ival

Pro

babi

lity

30 40 50

Residual CFTR activity (Class IV, V): R117H, R334W, R347P, 3849+10KbC–›T, 2789+5G–›A, A455E

Severely reduced CFTR activity (Class I, II, III): G542X, R553X, W1282X, R1162X, 621-IG–›T, 1717-1G–›A, 1078delT, 3659delC, I507del, N1303K, S549N, G85E, F508del, G551D, R560T

n=1126

n=14,525

• Life expectancy in Western countries (general population born in 2000) is ~79 years2

• Between 1993 and 2002, median survival for US patients with genotypes associated with little to no CFTR activity was 36.3 years (95% CI, 35.5 to 37.6 years), while median survival for those having genotypes associated with residual CFTR activity was 50 years (95% CI, 47.1 to 55.9 years)1

– In this study, patients with the 1717-1G–›A mutation (Class I) were part of the severely reduced CFTR activity group

• More recent US data (2000-2010) suggest median survival across genotypes continues to improve3

Survival Curves by CFTR Activity During a 10-Year Follow-Up (1993-2002) of Patients From the US CFF Registrya

a Data are from a retrospective study of patients enrolled in the Cystic Fibrosis Foundation patient registry measuring risk of death over a 10-year observation period from 1993 to 2002. Patients were grouped as having a high-risk or low-risk genotype based on the functional effects of their class of CFTR mutation on phenotype and mortality. Patients having a Class I, II, or III mutation on both alleles were considered high-risk, while patients having at least 1 Class IV or V mutation were categorized as low-risk. A total of 15,651 patients had a CFTR genotype of a known functional class; 14,525 (93%) had a high-risk CFTR genotype and 1126 (7%) had a low-risk CFTR genotype.1

This survival curve represents population-based outcomes.1 Individual outcomes in cystic fibrosis are variable.

Adapted with permission from McKone EF et al. Chest. 2006;130(5):1441-1447.

3

Country registries listing the 1717-1G–›A mutation report 0.1% to 3% prevalence among patients with CF1-10

• In the CFTR2 global database, ~1% of patients with CF have at least 1 copy of the 1717-1G–›A mutation11

Aus9: 2%

Brazil2: 0.1%

US1: 2%

Prevalence of the 1717-1G–›A Mutation in Patients With Cystic Fibrosis (% of Patients With at Least 1 Allele)

2%

2%

2%

2%

0.4%

Switzerland12

Italy13

Hungary13

Poland13

Canada14

Country % of Alleles

Additional sources report frequency of the 1717-1G–›A mutation on CF alleles

References: 1. Cystic Fibrosis Foundation. Cystic Fibrosis Foundation Patient Registry 2013 Annual Data Report. Bethesda, MD. © 2014; Cystic Fibrosis Foundation. 2. The Brazilian Cystic Fibrosis Study Group. Brazilian Cystic Fibrosis Patient Registry. 2012 Annual Report. 3. Belgisch Mucoviscidose Register – Registre Belge de la Mucoviscidose. The Belgian Cystic Fibrosis Registry. Summary Report 2011. © 2014; Institute of Public Health (WIV-ISP), Brussels, Belgium. 4. French Cystic Fibrosis Registry. Annual Report 2013. © Vaincre la Mucoviscidose and Ined, 2015; Paris, France. 5. Cystic Fibrosis Trust. UK Cystic Fibrosis Registry Annual Data Report 2014. © 2015; Cystic Fibrosis Trust; London, UK. 6. The Cystic Fibrosis Registry of Ireland. 2013 Annual Report. © CFRI, 2015; Dublin, Ireland. 7. Nederlandse Cystic Fibrosis Stichting. Dutch Cystic Fibrosis Patient Regristry. Report on the Year 2014. © 2015 NCFS. 8. Mukoviszidose e.V. und Mukoviszidose Institut gemeinnützige Gesselschaft für Forschung und Therapienntwicklung mbH. Beriichtsband Qualitätssicherung Mukoviszidose 2012. © 2013, Bonn, Germany. 9. Cystic Fibrosis Australia. Australian Cystic Fibrosis Data Registry 2013–16th Annual Report. © 2015; Cystic Fibrosis Australia; Baulkham Hills NSW, Australia. 10. PORT CFNZ National Data Registry, 2014 Registry Report, Cystic Fibrosis New Zealand. Christchurch, New Zealand. 11. US CF Foundation, Johns Hopkins University, The Hospital for Sick Children. The Clinical and Functional TRanslation of CFTR (CFTR2). http://www.cftr2.org. Accessed November 12, 2015. 12. European Cystic Fibrosis Society. ECFS Patient Registry 2010 Annual Data Report. 2014. 13. Bobadilla JL et al. Hum Mutat. 2002;19(2):575-606. 14. World Health Organization. The molecular genetic epidemiology of cystic fibrosis: report of a joint meeting of WHO/ECFTN/ICF(M)A/ECFS; June 19, 2002; Genoa, Italy. © World Health Organization; 2004.

Europe:Belgium3: 3%France4: 2% UK5: 1%Ireland6: 1%Netherlands7: 1% Germany8: 0.7%

New Zealand10: 0.5%

4

The 1717-1G–›A mutation results in defective biosynthesis of the CFTR protein1-3

References: 1. Hull J et al. Hum Mol Genet. 1993;2(6):689-692. 2. Green DM et al. Respir Res. 2010;11:140. doi: 10.1186/1465-9921-11-140. 3. Zielenski J. Respiration. 2000;67(2):117-133.

• 1717-1G–›A is a splice mutation, which produces a premature stop codon1

• The cell cannot synthesize a full-length CFTR protein, a Class I mutation1-3

• As a result, few to no CFTR proteins are present at the apical cell surface3

CFTR Processing

CFTR Synthesis

CFTR Trafficking

Endoplasmicreticulum

Proteosome

Golgicomplex

T

T

T

T

T

T

A

A

A

G

C

C

C

C

C

Rest ofCFTRgene

CFTR gene(DNA)

EndoplasmicReticulum

mRNA

DNA

CFTR gene

mRNA Pre-mRNA

RNA processing

Splicingdefect

Chromosome 7

DNA

Transcription

Illustrative Example of Class I Defect

CFTR Turnover

CFTR Function

5

The 1717-1G–›A allele results in little to no total CFTR activity1-5

Defective Synthesis (Class I)

1717-1G–›A allele results in few to no CFTR channels at

apical surface

Channel-open Probability: N/A Conductance: N/A

Little to No1717-1G–›A–CFTR

Activity

CFTR Function

A virtual absence of 1717-1G–›A-CFTR protein quantity…

…regardless of function since few to no CFTR proteins reach the surface…

…results in little to no total CFTR activity

References: 1. Hull J et al. Hum Mol Genet. 1993;2(6):689-692. 2. Green DM et al. Respir Res. 2010;11:14. doi: 10.1186/1465-9921-11-140. 3. Zielenski J. Respiration. 2000;67(2):117-133. 4. Welsh MJ, Smith AE. Cell. 1993;73(7):1251-1254. 5. Sheppard DN et al. Nature. 1993;362(3):160-164.

1

1

2

2

3

3

Total CFTR activity can be defined as total ion transport mediated by CFTR protein channels at the cell surface, depending on CFTR protein quantity and function.5

x x =

Channel-openProbability

Conductance

CFTR FunctionTotal

CFTR ActivityxCFTR Quantity x =

N/A, not applicable.

6

Both CFTR alleles play a role in determining phenotype or disease severity1-6

• A 1717-1G–›A allele results in little to no CFTR activity. The phenotype of a particular patient is also influenced by the mutation on the other allele1-6

• 1717-1G–›A typically results in the indicated phenotypes

No CF DiseaseNormal individuals and CF carriers

Cystic Fibrosis

CFTR-related DisorderClinical entities associated with CFTR dysfunction that do not fulfill diagnostic criteria for CF

Normal Normal Normal

Normal Residual

Residual Residual

Residual

Littleto None

Littleto None

Littleto None

Littleto None

Allele 1Total CFTR Activity

Allele 2Total CFTR Activity

TotalCFTR

Activity

0%

100%

References: 1. US CF Foundation, Johns Hopkins University, The Hospital for Sick Children. The Clinical and Functional TRanslation of CFTR (CFTR2). http://www.cftr2.org. Accessed November 12, 2015. 2. deGracia J et al. Thorax. 2005;60(7):558-563. 3. Cystic Fibrosis Genotype-Phenotype Consortium. N Engl J Med. 1993;329(18):1308-1313. 4. Davis PB et al. Am J Respir Crit Care Med. 1996;154(5):1229-1256. 5. Castellani C et al. J Cyst Fibros. 2008;7(3):179-196. 6. Zielenski J. Respiration. 2000;67(2):117-133.

Adapted from Zielenski J. Respiration. 2000;67(2): 117-133.

7

1717-1G–›A in combination with another allele that produces little to no CFTR activity usually results in a CF phenotype1-5

References: 1. US CF Foundation, Johns Hopkins University, The Hospital for Sick Children. The Clinical and Functional TRanslation of CFTR (CFTR2). http://www.cftr2.org. Accessed Novemebr 12, 2015. 2. deGracia J et al. Thorax. 2005;60(7): 558-563. 3. Cystic Fibrosis Genotype-Phenotype Consortium. N Engl J Med. 1993;329(18):1308-1313. 4. Davis PB et al. Am J Respir Crit Care Med. 1996;154(5):1229-1256. 5. Castellani C et al. J Cyst Fibros. 2008;7(3):179-196.

CF Phenotype In patients registered in the CFTR2 database with a 1717-1G–›A mutation on 1 allele and a pancreatic insufficient mutation on the second allele1

• Elevated sweat chloride (average):103 mmol/L

• Lung function decline over time

• Pseudomonas colonization: 54% of patients

• Pancreatic insufficiency: 97% of patients

Little to NoTotal CFTR

Activity

EnvironmentalFactors

Modifier Genes

Little to NoCFTR Protein

Activity

CFTR Genotype

Allele #1: 1717-1G–›A

Little to NoCFTR Protein

Activity

Allele #2

8

Summary• Loss of CFTR activity is the underlying cause of CF

• Levels of CFTR activity affect survival in CF

• Country registries listing the 1717-1G–›A mutation report 0.1% to 3% prevalence among patients with CF

• The 1717-1G–›A mutation results in defective biosynthesis of the CFTR protein

• The 1717-1G–›A allele results in little to no total CFTR activity

• Both CFTR alleles play a role in determining phenotype or disease severity

• 1717-1G–›A in combination with another allele that produces little to no CFTR activity usually results in a CF phenotype

9